The objective of the proposed research is to understand at the molecular level the genetic regulation of cellular and flagellar differentiation in the bacterium Proteus. From laboratory studies, it is apparent that the same strain of Proteus can express one phenotype appropriate for growth in liquid media and another phenotype appropriate for growth on surfaces. When grown in liquid media, Proteus produce very short rods about 0.6 mu(m) wide and 1-2 mu(m) long. These cells possess between 1 and 10 peritrichously arranged flagella per bacterium. When cells are propagated on media solidified with agar, the bacteria undergo a dramatic morphological change. Shortly after encountering a solid surface, cellular division stops and the bacteria begin to elongate. This process can result in an elongated, multinucleated "swarmer" cell 20 to 80 mu(m) in length, and occurs concomitantly with the synthesis of new flagellar structures. While the short liquid-grown cells possess few flagella, estimates of the number of flagella per swarmer cell range from 500 to 5000. If the swarmer cells are transferred from an agar medium to a liquid medium, they start to septate and divide. This process of de- differentiation results in a cellular morphology typical of broth-grown cells. Mutational analyses of Proteus using transposon mutagenesis to transcriptionally couple the genes responsible for differentiation (swr) to reporter genes, such as lux and lacZ, will be undertaken. Swr- mutants will be identified from a bank of transductants, screened for surface- induced gene activity, and grouped into phenotypically related categories. We will attempt to identify the stimulus evoking swr gene expression. These efforts will be guided by our previous data showing that physical stimulus (viscosity) is the crucial factor influencing swr gene expression. We will also analyze the kinetics of swr gene activity when cells are placed under inducing conditions, and when the inducer is removed. Such data will help establish a regulatory hierarchy of the swr gene control. Transposon insertions in the regulatory locus (or loci) will be cloned from Proteus into E. coli and analyzed using recombinant DNA methodologies. The results stemming from this project will help us understand how cell sense their physical environment and how that stimulus controls the expression of genes involved in cellular differentiation.